KR20010029658A - Agent for enhancing cerebral acetylcholine release - Google Patents

Abstract

PURPOSE: Provided is a pharmaceutical composition for increasing cerebral acetylcholine release. Particularly, it comprises N-anisoyl-gamma-aminobutyric acid or p-anisic acid and is used in treating circadian rhythm, sleep and attention deficit disorders. CONSTITUTION: Pharmaceutical composition for increasing cerebral acetylcholine release comprises N-anisoyl- psi -aminobutyric acid or p-anisic acid and an therapeutically inert carrier, as active ingredients. The effective dose of N-anisoyl- psi -aminobutyric acid or p-anisic acid is preferably 1- 300mg/kg per day.

Description

Acetylcholine release enhancer in the brain {AGENT FOR ENHANCING CEREBRAL ACETYLCHOLINE RELEASE}

The present invention relates to the usefulness of N-anisoyl-γ-aminobutyric acid (N-anisoyl-GABA) or p-anisic acid as a pharmaceutical composition that enhances acetylcholine (ACh) release in the brain. In particular, the present invention relates to pharmaceutical compositions for circadian rhythm disorders, sleep disorders, attention deficit disorders and problematic behaviors.

ACh has been widely known as an important typical neurotransmitter. Numerous cholinergic pathways exist in the nucleus-thalamus, septal-hippocampus and basal-nucleus-neocortex of mesopontin reticular tissue in the brain of mammals (Neuroscience, 10, 1185-1201 (1983)). This cholinergic pathway has been taught to play a pivotal role in the induction and maintenance of rapid eye movement sleep (REMS) and in the regulation of arousal and attention, learning and memory, and drivers [Journal of Neuroscience, 10, 2541-2559 ( 1990); Brain Research Review, 19, 298-318 (1994). However, patients suffering from neuronal degeneration (ie Alzheimer's disease, Parkinson's disease and advanced nuclear palsy) [Lancet, 2, No. 8000, 1403 (1976); Journal of Neurological Neurosurgical Psychiatry, 51, 540-543 (1988) and patients suffering from cerebrovascular disease [Dementia, 5, 163-167 (1994); Journal of Neural Transmisson, 103, 1211-1220 (1996)] decreased central cholinergic neuronal activity. It has been suggested that the cholinergic deficiency and / or dysfunction is associated with a variety of neuropsychiatric symptoms such as dementia, sleep disorders, low awakening, lack of attention and delirium, and problematic behaviors including nighttime wandering.

1-p-anisoyl-2-pyrrolidinone (anisetam, European Patent Nos. 5143 and 44088), a brain function improving agent, is used for the treatment of emotional disorders (anxiety / earness and depressed mood) that appear as sequelae after cerebral infarction. Igaku No Ayumi, 156, No. 2, 143-187 (1991); Geriatric Medicine, 36, 1513-1520 (1998). Although it has been reported that the drug activates the central cholinergic system, it is not clear whether the substance (s) comprising its metabolites cause significant cholinergic activity by orally administered 1-p-anisoyl-2-pyrrolidinone. Drug Investigation, 5, 1-108 (1993). Moreover, the biological activities of N-anisoyl-GABA and p-anisic acid, the major in vivo metabolites of 1-p-anisoyl-2-pyrrolidinone, are not clearly explained.

On the other hand, treatment of the aforementioned diseases is initiated by increasing central ACh levels using ACh esterase inhibitors. Because ACh esterase is widely distributed not only in the brain but also in peripheral tissues, peripheral side effects caused by adverse blood-brain barrier penetration of ACh esterase inhibitors can cause serious problems for these enzyme inhibitors as drugs. Thus, the present invention provides a preferred compound that can safely increase central ACh levels to address this problem.

It was surprisingly found that N-anisoyl-GABA and p-anisic acid respectively enhance brain ACh release in the brain.

Accordingly, the present invention relates to the use of N-anisoyl-GABA or p-anisic acid, preferably in the form of pharmaceutical compositions, to enhance ACh release in mammalian, preferably human brain. N-anisoyl-GABA or p-anis acid compounds and pharmaceutical compositions comprising the same are particularly useful for the treatment of circadian rhythm disorders, sleep disorders, attention deficit disorders and problematic behaviors.

The present invention relates to a pharmaceutical composition for enhancing the release of acetylcholine in the brain comprising N-anisoyl-GABA or p-anisic acid as an active ingredient for the treatment of the disease.

The invention also relates to methods of using N-anisoyl-GABA or p-anisic acid to enhance ACh release in the brain, particularly for the treatment of circadian rhythm disorders, sleep disorders, attention deficit disorders and problem behaviors. .

The invention is explained in detail below. The following description and examples are intended to aid the understanding of the present invention, but should not be construed as limiting the invention in this way. The results obtained in the examples are demonstrated by the accompanying drawings.

FIG. 1 shows anisocetam (A), 2-pyrrolidinone, p-anisane and N-anisoyl-GABA (B) for ACh release in the reticulum nucleus of thalamic spontaneous hypertensive rats (SHRSP) of freely moving seizures. ) Shows the effect. The compound was perfused for 20 minutes (solid bar). Data represent mean ± SEM. Baseline ACh release was 0.35 ± 0.03 mmol / 20 min (n = 24). ^* P <0.05 compared to vehicle control.

FIG. 2 shows the effects of anicetam (A), 2-pyrrolidinone, p-anisane and N-anisoyl-GABA (B) on ACh release in the posterior hippocampus of freely moving SHRSP. The compound was perfused for 20 minutes (solid bar). Data represent mean ± SEM. Baseline ACh release was 0.63 ± 0.04 mmol / 20 min (n = 15). In comparison to setam, ^* P <0.05, ^** P <0.01.

FIG. 3 shows the effects of butasetam (A), 2-pyrrolidinone, p-anisane (B) and N-anisoyl-GABA (C) on ACh release in the cortex of the frontal lobe of freely moving SHRSP. . The compound was perfused for 20 minutes (solid bar). Data represent mean ± SEM. Baseline ACh release was 0.58 ± 0.03 mmol / 20 min (n = 24). In comparison to setam, ^* P <0.05, ^** P <0.01.

4 shows the predicted activity associated with circadian motility activity rhythms and mealtimes in young and old rats. After fasting for 24 hours (the same day), feeding was limited to only 1 hour from 13:30 for 6 consecutive days (1 to 6 days). The seventh day was not fed again. Data were obtained from seven rats per group and represent the mean ± S.E.M. Of motor activity measured every hour. Dotted pillars indicate the feeding period. Δ is -1 day; ○ is 6 days; ● is 7 days.

FIG. 5 shows the effect of butestam on anticipated activity rhythms associated with circadian motility and mealtime in old rats. Rats were orally administered to rats immediately after feeding once daily for 7 consecutive days (1 to 7 days). Data represent mean ± S.E.M. Obtained from 5 to 6 rats per group. Dotted pillars indicate the feeding period. ○ is 6 days; ● is 7 days.

FIG. 6 depicts daytime and nighttime changes of Wistar Kyoto rats (WKY) associated with REMS, NREMS, and brain temperature and age in SHRSP. The data represent the mean ± S.E.M. Of each of the measured parameters every hour.

FIG. 7 shows the effect of repeated administration of butamstam on REMS, NREMS, and brain temperature in SHRSP. Animals were orally administered noncetamam twice daily (09:00 and 20:00) for 5 consecutive days and data after 9 and 10 doses are shown.

N-anisoyl-GABA and p-anisic acid are both known drug compounds. N-anisoyl-GABA can be synthesized by the method described in Spanish Patent Publication No. 84-538772. P-anisic acid can be prepared by the method described in Journal of American Chemical Society 78,907-909 (1956), and Sigma Chem. Sigma Chem. Co., St. Louis, USA, Lancaster Synthesis Ltd., Lancashire, UK, Waco Pure Chem. It is also available from Wako Pure Chem. Ind. Ltd. (Osaka, Japan) and the like.

N-anisoyl-GABA and p-anisic acid may each be used in the form of a pharmaceutically acceptable formulation. Such formulations may be in the form of tablets, coated tablets, sugars, hard gelatin capsules, soft gelatin capsules, as well as solutions, emulsions or suspensions. Such forms can be administered orally. In addition, the preparations may be in the form of suppositories for rectal administration or in the form of injections which may be administered parenterally.

When preparing oral solid preparations such as tablets, coated tablets, sugars, or hard gelatin capsules, N-anisoyl-GABA or p-anisic acid, respectively, may be combined with lactose, maize or corn starch and derivatives thereof. Such pharmaceutically inert inorganic or organic carriers, talc, stearic acid and bases or salts thereof, and the like.

When forming soft or hard gelatin capsule products, carriers such as, for example, vegetable oils, waxes, fats, oils, gels, semisolid or liquid polyols may be suitably used.

When forming liquid and syrup products, carriers such as, for example, water, polyols, sucrose, invert sugar, glucose and the like may suitably be used.

When forming an injectable product, carriers such as, for example, water, alcohols, polyols, glycerol, vegetable oils can be suitably used.

When forming suppositories, carriers such as, for example, vegetable oils, waxes, oils, gels or liquid polyols may be suitably used. In addition, such formulations may be used in admixture with preservatives, solvents, stabilizers, wetting agents, emulsifiers, sweeteners, bases that change the osmotic pressure, buffers, encapsulations and antioxidants, and further therapeutically noteworthy compounds.

The route of administration of the formulation is not limited to the above, and may be appropriately changed depending on the form of the formulation or the age, sex, and symptoms of the patient.

Routes of administration, dosages, and multiple administrations may be appropriately modified depending on age, weight, and symptoms of the patient. For oral administration, the daily dose for normal adults is 1 to 300 mg / kg (preferably 3 to 30 mg / kg) and such doses may be administered from one to several times.

When orally administered N-anisoyl-GABA or p-anisic acid to rats, the acute toxicity (LD ₅₀ value) of N-anisoyl-GABA is greater than 5,000 mg / kg in both males and females, and p-anisic acid The acute toxicity of was 1,813 and 2,124 mg / kg in males and females, respectively. Subsequent oral administration of both of these drugs to rats over 4 weeks showed no subacute toxicity, such as mortality and malformations, below 600 mg / kg in hematology, blood biochemistry and toxicology tests.

The effect of enhancing ACh release by N-anisoyl-GABA and p-anisic acid and the improvement of circadian rhythm disorder and sleep disorder by 1-p-anisoyl-2-pyrrolidinone are described in detail below.

Example 1

Test for ACh Release Enhancer Effect

Test Animals: Male 13-week-old spontaneous hypertensive rats (SHRSP) males who received 1% NaCl solution instead of water for 5 weeks before the experiment.

Test Methods: SHRSP was anesthetized and a ceramic cannula was inserted into the cortex of the thalamus, posterior hippocampus and frontal lobe. After recovery, a concentric microseparation probe was inserted into the ceramic cannula and under normal conditions of freely moving conditions, a normal ringer containing 10 ^-5 M of serine (SIGMA, St. Louis, USA) at a constant flow rate of 2 μL / min. Perfused with solution. N-anisoyl-GABA or p-anisic acid was dissolved in the Ringer's solution at final concentrations of 10 ⁻⁷ , 10 ⁻⁶ and 10 ⁻⁵ M and each drug was perfused for 20 minutes through the same probe. Dialysis was collected every 20 minutes and inserted into a high pressure liquid chromatography system to quantify extracellular ACh levels. ACh release is expressed as percent change relative to the mean of three continuous stable samples collected before perfusion of the drug.

Test results: N-anisoyl-GABA increases ACh release to 32%, 48% and 70%, p-anisic acid increases ACh release, thalamus (Figure 1), posterior hippocampus (Figure 2) and the frontal cortex ( Increased to 22%, 51% and 61%, respectively. In contrast, the effect of 1-p-anisoyl-2-pyrrolidinone was not observed in any brain region. Thus, N-anisoyl-GABA and p-anisic acid were considered to be active substances that contribute to the activity of central cholinergic neurons (ACh release enhancement) as the main metabolites of 1-p-anisoyl-2-pyrrolidinone.

Example 2

Tests for Impaired Diurnal Rhythm Improvement

Test animals: Male Wistar rats at 9 weeks (young group) and 30 months (old group).

Test Method: Animals were kept in cages each free to use food and water. After fasting for 24 hours (same day), the animals were fed with restriction from 13:30 to 1 hour per day for 6 consecutive days. Rather than feeding again on the seventh day, the predicted circadian activity induced by feeding was examined. 30- and 100 mg / kg of 1-p-anisoyl-2-pyrrolidinone or vehicle was administered orally immediately after the feeding time once daily for 7 consecutive days. Spontaneous motor activity was measured in each cage.

Test Results: Expected activity induced by feeding at 7 days in old rats was significantly reduced compared to young rats, abnormal in circadian rhythm control, and lacking ability to keep time with age. Repeated oral administration of 1-p-anisoyl-2-pyrrolidinone, except vehicle, ameliorated the anticipated activity worsened in old rats (FIG. 5).

Example 3

Test for improving anxious sleep patterns

Test Animals: 13 week old SHRSP males and 1 year old Wistar Kyoto rats (WKY) who consumed 1% NaCl solution instead of water over 5 weeks.

Test Methods: Animals were anesthetized, electrodes for EEG and sensors for brain temperature were inserted into the cortex of the brain, and electrodes for EMG (EMG) were inserted into the back cervical muscles. All variables were recorded continuously for 7 days and the amplitude and frequency of the EEG and EMG waves were analyzed to classify the rat's active state into awake state, REMS and NREMS. 15 mg / kg of 1-p-anisoyl-2-pyrrolidinone or vehicle was administered orally twice daily (morning and evening) for 5 consecutive days.

Test Results: Compared with the comparative WKY, SHRSP showed a decrease in REMS during daytime (sleep period), an increase in NREMS during nighttime (activity period) and a decrease in brain temperature (Figure 6). . Repeated oral administration of 1-p-anisoyl-2-pyrrolidinone except vehicle improved the reduced weekly REMS in SHRSP (FIG. 7).

Based on the above, circadian rhythms observed in cerebrovascular disease (ie cerebral infarction and cerebral hemorrhage) as well as neuronal degeneration (ie Alzheimer's disease, Parkinson's disease and progressive nuclear palsy) and hyperkinetic symptoms (attentional hyperactivity) It can be concluded that N-anisoyl-GABA or p-anisane is useful for the treatment of various neuropsychiatric symptoms such as disorders, sleep disorders, attention deficit disorders and problematic behaviors (delivers and nighttime wandering). .

Example 4

Preparation of pharmaceutical composition

The pharmaceutically acceptable formulations provided below are merely suitable examples and should not be construed as limiting the pharmaceutical composition of the invention in any way.

4.1 Preparation of Tablets Containing N-Anisoyl-GABA

A tablet containing 100 mg of N-anisoyl-GABA was prepared by the following method using the following composition (tablet sugar).

Composition (A):

N-anisoyl-GABA 100mg

Lactose 20mg

Kollidon CL (BASF) 15 mg

Corn starch 30mg

Avicel PH 101 (Asahi Chemical Co., Ltd) 50 mg

Composition (B):

Polyvinylpyrrolidinone K-90 5mg

Photosilicate anhydrous 18mg

Magnesium Stearate 2mg

240 mg in total

The mixture of composition (A) described above is mixed in 8% aqueous polyvinylpyrrolidinone K-90 solution. It is dried at 60 ° C. and then mixed with composition (B). This mixture is tableted into round tablets weighing 240 mg and 8 mm in diameter.

4.2 Preparation of tablets containing p-anisic acid

A tablet containing 100 mg of p-anisic acid was prepared by the following method using the following composition (tablet sugar).

Composition (A):

100 mg of p-anisic acid

Lactose 20mg

Collidone CL (BASF) 15 mg

Corn starch 30mg

Avicel PH 101 (Asahi Chemical Company Limited) 50 mg

Composition (B):

Polyvinylpyrrolidinone K-90 5mg

Photosilicate anhydrous 18mg

Magnesium Stearate 2mg

240 mg in total

The mixture of composition (A) is mixed in 8% aqueous polyvinylpyrrolidinone K-90 solution. It is dried at 60 ° C. and then mixed with composition (B). The mixture is tableted into round tablets weighing 240 mg and 8 mm in diameter.

4.3 Preparation of Capsules Containing N-Anisoyl-GABA

A capsule containing 100 mg of N-anisoyl-GABA was prepared by the following method using the following composition (per capsule).

Composition (A):

N-anisoyl-GABA 100mg

Lactose 20mg

Collidone CL (BASF) 2mg

Corn Starch 53mg

Composition (B):

Polyvinylpyrrolidinone K-90 5mg

Avicel PH 101 (Asahi Chemical Company, Limited) 18mg

Magnesium Stearate 2mg

200 mg in total

The mixture of composition (A) described above is mixed in 8% aqueous polyvinylpyrrolidinone K-90 solution. It was dried at 60 ° C. and then mixed with composition (B). This mixture is poured into gelatin capsules three times to give a capsule containing 200 mg.

Pharmaceutical compositions comprising N-anisoyl-GABA and p-anisic acid according to the present invention are used to safely increase ACh release in the brain and improve circadian rhythm disorders, sleep disorders, attention deficit disorders and problematic behaviors.

Claims (3)

A pharmaceutical composition for enhancing acetylcholine release in the brain, comprising N-anisoyl-GABA or p-anisic acid as a active ingredient and a therapeutically inert carrier. The method of claim 1, Pharmaceutical compositions for the treatment of circadian rhythm disorders, sleep disorders, attention deficit disorders and problem behaviors. The method according to claim 1 or 2, A pharmaceutical composition wherein the active ingredient N-anisoyl-GABA or p-anisic acid is administered to an adult at a dose of 1 to 300 mg / kg.